Structural and thermodynamic insights into β-1,2-glucooligosaccharide capture by a solute-binding protein in Listeria innocua

β-1,2-Glucans are bacterial carbohydrates that exist in cyclic or linear forms and play an important role in infections and symbioses involving Gram-negative bacteria. Although several β-1,2-glucan-associated enzymes have been characterized, little is known about how β-1,2-glucan and its shorter oligosaccharides (Sopn_s) are captured and imported into the bacterial cell. Here, we report the biochemical and structural characteristics of Sopn-binding protein (SO-BP, Lin1841) associated with the ABC transporter from the Gram-positive bacterium Listeria innocua. Calorimetric analysis revealed that SO-BP specifically binds to Sopn_s with degree of polymerization of 3 or more, with Kd values in the micromolar range. The crystal structures of SO-BP in an unliganded open form and in closed complexes with tri-, tetra-, and pentaoligosaccharides (Sopn_3-5) were determined to a maximum resolution of 1.6 Å. The binding site displayed shape complementarity to Sopn,which adopted a zigzag conformation. We noted that water-mediated hydrogen bonds and stacking interactions play a pivotal role in the recognition of Sopn_3-5by SO-BP, consistent with its binding thermodynamics. Computational free-energy calculations and a mutational analysis confirmed that interactions with the third glucose moiety of Sopn_s are significantly responsible for ligand binding. A reduction in unfavorable changes in binding entropy that were in proportion to the lengths of the Sopn_s were explained by conformational entropy changes. Phylogenetic and sequence analyses indicated that SO-BP ABC transporter homologs, glycoside hydrolases, and other related proteins are co-localized in the genomes of several bacteria. This study may improve our understanding of bacterial β-1,2-glucan metabolism and promote discovery of unidentified β-1,2-glucan-associated proteins.